The present application relates to combinations of a heparin cofactor II agonist and a platelet glycoprotein IIb/IIIa receptor (GPIIb/IIIa) antagonist that are useful in inhibiting both platelet aggregation and thrombin generation resulting from disease, or injury responses to wound repairs. The present application particularly relates to the use of subtherapeutic amounts of a heparin cofactor II agonist and subtherapeutic amounts of a platelet GPIIb/IIIa receptor antagonist that, in combination, are therapeutically effective in inhibiting both platelet aggregation and thrombin generation.
Cardiovascular disease is the primary cause of death in the USA. According to the American Heart Association, 2.5 million individuals suffer from venous thrombosis and 600,000 suffer from pulmonary embolism each year. In 1996, approximately 830,000 cardiac surgeries and 700,000 cardiac catheterization procedures were performed in the USA as a result of arterial and venous thromboses. Usually, anticoagulant therapy is implemented either alone or in combination with anti-platelet and/or anti-fibrinolytic therapies, particularly in acute care settings where the immediate reopening of a blocked vessel becomes imperative. The drugs used in these therapies, however, have certain dose-limiting side effects, the foremost being hemorrhagic (i.e., prolonged bleeding) and when used in combination, these side effects can become potentiated, further limiting effective dosing and duration of the needed drug treatment. See Fareed, xe2x80x9cDrug Interactions with Antiplatelet Agentsxe2x80x9d IBC 3rd Annual Mini-Symposium on Advances in Antiplatelet Therapies (Waltham, Mass. 2000).
With current anticoagulants, the bleeding effects are due to an action on one or more of the enzymes that regulate hemostasis in the global circulation, versus their action in a more specific and limited sense on enzymes of the hemostatic mechanism that promote the disease process at the vascular wall, e.g., low selectivity. Likewise, antiplatelet drugs exhibit strong interactions with the anticoagulants (such as heparin), antithrombin drugs and thrombolytic agents, and safety considerations, for example, preclude their administration to patients at high risk for intracranial hemorrhage, particularly elderly patients with poorly controlled hypertension and previous manifestations of cerebrovascular disease.
Central to this problem is control of thrombin generation and activity. This enzyme plays a key role in the formation of venous and arterial occlusions and in the causation of platelet emboli. Also key to this problem is achieving the sustained inhibition of thrombin at the diseased site which otherwise perpetuates its continued generation in an unabated fashion through a thrombin feedback mechanism that drives clot growth and platelet activation. A more targeted inhibition of thrombin at the disease site and the platelet surface using agents or drugs of higher selectivity would cause fewer side effects on the blood coagulation properties of the peripheral circulation and thus potentially allow safer and more effective dosing regimens in combination therapies.
Processes that compromise the integrity of the vascular wall result in the activation of the hemostatic mechanism affecting the blood coagulation cascade and platelet activation pathways. See Furie et al., xe2x80x9cMolecular and Cellular Biology of Blood Coagulation,xe2x80x9d N. Eng. J Med (1992) 326: 800-806. This response to wound repair results in the growth of a thrombus forming an occlusion that impedes the flow of blood and thus oxygen and needed nutrients to the vital tissues. For example, atherosclerosis is a disease process affecting the coronary arteries and major arterioles of the heart in which both inflammatory reactions (leukocytes, neutrophils, complement activation) and the accumulation of lipids (e.g., cholesterol, cholesterol esters, saturated fats, oxidized lipids and foam cells) occur. These events are toxic to the endothelial cells that line the blood vessel wall, the purpose of these cells being to form a protective non-thrombogenic surface or barrier separating blood from tissue. The exfoliation of the endothelial cells exposes blood to the subendothelial surface which has a high thrombogenic potential. This results in the activation of the blood coagulation cascade and the generation of active thrombin. This active thrombin becomes bound to the disease site and promotes the formation of the clot. Contact of blood with foreign surfaces such as those of extracorporeal circuits and vascular devices (stents, guidewires, etc.) also induces thrombin generation.
Thrombin converts soluble fibrinogen into insoluble fibrin at the vascular injury site where it is stabilized by enzymatic crosslinking reactions and platelet interactions. Thrombin is a potent platelet agonist and can interact on the platelet surface with receptors that lead to activation. See Furman et al, xe2x80x9cThe Cleaved Peptide of the Thrombin Receptor Is a Strong Platelet Agonist,xe2x80x9d Proc. Natl. Acad Sci. (1998) 95(6):3082-3087; Zucker et al, Platelet Activation Arteriosclerosis (1985) 5(1):2-18. This leads to a thrombus rich in fibrin and platelets that may then become occlusive to the flow of blood to the heart and other organs such as the brain, resulting in serious life-threatening illnesses such as myocardial infarction and stroke.
There are many variations of vessel disease of the arterial and venous circulations. Clots of the arterial side tend to be enriched in platelets whereas those on the venous side contain fewer platelets and are enriched in fibrin. Thrombo-embolic diseases involving thrombus formation of the arterial and venous circulations include acute coronary syndromes (ACS), myocardial infarction (MI), deep vein thrombosis (DVT), pulmonary embolism (PE) and stroke to name a few. Procedures involving clamping of arteries such as carotid endarterectomy and peripheral vascular surgery also induce vascular damage, thrombin formation and platelet activation. Invasive cardiovascular procedures such as coronary artery bypass grafts (CABG), percutaneous transluminal coronary angioplasty (PTCA), cardiac catheterizations and the use of extracorporeal interventions, including cardiopulmonary bypass surgery (CPB), end-stage renal dialysis (ESRD) and extracorporeal membrane oxygenation (ECMO), potently activate the clotting system and affect platelet function.
Heparin-induced thrombocytopenia (HIT) is a special class of platelet thrombosis that occurs as an immune response to heparin, the anticoagulant drug most often employed first in the prevention and treatment of thrombo-embolic diseases. HIT leads to a precipitous drop in platelet count, an increase in platelet-induced thrombin generation and potentially to a fatal thrombosis. Standard treatment of HIT involves the discontinuation of heparin and use of an alternative anticoagulant such as a thrombin inhibitor, followed by close patient monitoring for the recovery of platelet counts. Despite the use of these alternatives, the morbidity and mortality of HIT patients remains high. Recently, a standard dose of GPIIb/IIIa antagonist, combined with a lowered dose of thrombin inhibitor to minimize hemorrhagic events, was used to treat HIT thrombosis. See Walenga et al, xe2x80x9cClinical Experience with Combined Treatment of Thrombin Inhibitors and GPIIb/IIIa Inhibitors in Patients with HIT,xe2x80x9d Semin. Thromb. Hemost. (1999) 25 (suppl. 1):77-81. While initial thrombosis of the coronary arteries tends to be susceptible to first treatment with fibrin-dissolving agents (e.g., tissue plasminogen activator or streptokinase), a fibrinolytic-resistant re-thrombosis often occurs that is platelet-rich. This most often requires the use of fast-acting antiplatelet drugs such as GPIIb/IIIa antagonists combined with thrombin inhibitors to control the local generation of active thrombin. However, more effective combinations of improved anticoagulants in combination with the GPIIb/IIIa antagonists are needed in the treatment of HIT and other thrombo-embolic disorders.
These improved anticoagulants require greater selectivity for thrombin at the diseased site. Surface-bound thrombin at residual levels amplifies the generation of systemic thrombin by catalyzing prothrombin consumption via the thrombin feedback loop at the site of vascular injury. See, for example, Ofosu et al, xe2x80x9cThrombin-Catalyzed Amplification and Inhibitory Reactions of Blood Coagulation in Thrombin: Its Key Role in Thrombogenesis-Implications for its Inhibition Clinically,xe2x80x9d CRC Press (1995) pp. 1-18. Moreover, when thrombin is generated in response to an injury or disease, it can be found not only in the systemic circulation or fluid phase, but is also associated with the fibrin clot, with cell surfaces such as platelets, the vessel wall and with the biomaterial surfaces of biometric circuits and devices.
Heparin affects the potent inhibition of systemic thrombin and is widely effective in the management of these thrombotic states. However, it is relatively ineffective in bringing about the inhibition of surface-bound thrombin key to the self-promotion of systemic thrombin generation. Evidence suggests that heparin may enhance clot growth when bound to the clot. See Kumar et al, xe2x80x9cThe Influence of Fibrinogen and Fibrin on Thrombin Generation-Evidence for Feedback Activation of the Clotting System by Clot Bound Heparin,xe2x80x9d Thromb. Hemost. (1994) 72: 713-721. Heparin""s principle mode of action occurs at the level of antithrombin III (AT), a circulating proteinase inhibitor that binds thrombin and other factors of the coagulation cascade to block their activity. Heparin serves as a template to promote the assembly of the thrombin-antithrombin III complex (TAT) that then binds to exosite 2 on the surface of systemic thrombin, thereby forming a ternary complex which greatly accelerates the second order rate constant for thrombin inhibition by the serine proteinase inhibitor. However, when thrombin becomes surface-bound, such as to the fibrin clot, exosite 2 on the thrombin surface becomes unavailable to the HAT complex and surface-bound thrombin resists inhibition. Thus, recurrent thrombosis may ensue following the discontinuation of heparin therapy. See Hogg et al, xe2x80x9cFibrin Monomer Protects Thrombin from Inactivation by Heparin-Antithrombin III: Implications for Heparin Efficacy,xe2x80x9d Proc. Natl. Acad. Sci. U.S.A. (1989) 86:3619-23.
European patent application 668,875 and PCT application WO 94/09034A 1 disclose a targeted-anticoagulant concept where the efficacy of heparin to inhibit clot bound thrombin is increased by its covalent attachment to a fibrin-specific monoclonal antibody used to deliver the glycosaminoglycan (or drug) into the clot. The efficacy of this approach with respect to heparin is still limited by the unavailability of exosite 2 on clot-bound thrombin that is critical to the binding of the HAT complex. Moreover, such targeted-anticoagulant concepts do not address the catalytic thrombins that remain active and bound to surfaces such as the platelet membrane, vessel wall or biomaterial surfaces of extracorporeal circuits. Even with these targeted-anticoagulant concepts, thrombin generation can be perpetuated at other sites, causing the disease process to linger.
Clot-bound heparin is susceptible to inhibition by exosite 1 directed inhibitors such as the leech anticoagulant peptide hirudin and heparin cofactor II. See Weitz et al, xe2x80x9cClot-Bound Thrombin Is Protected from Inhibition by Heparin-Antithrombin III But Is Susceptible to Inactivation by Antithrombin III-Independent Inhibitors,xe2x80x9d J. Clin. Invest. (1990) 86: 385-391; Bendayan et al., xe2x80x9cDermatan Sulfate is a More Potent Inhibitor of Clot-Bound Thrombin Than Unfractionated and Low Molecular Weight Heparins,xe2x80x9d Thromb. Haemost. (1994) 71:576-580. However, like heparin, hirudin exhibits significant bleeding side effects associated with its use . See Kwapis et al, xe2x80x9cProlonged Bleeding After Cardiopulmonary Bypass with Recombinant Hirudin,xe2x80x9d Eur. J. Cardiothorac. Surg. (1999) 16(2):256-257; Gast et al, xe2x80x9cInhibition of Clot-Bound and Free (Fluid-Phase Thrombin) by a Novel Synthetic Thrombin Inhibitor (Ro 46-6240), Recombinant Hirudin and Heparin in Human Plasma,xe2x80x9d Blood Coagul. Fibrinolysis (1994) 5(6):879-887. Although hirudin has a marginally increased selectivity for clot-bound heparin, fluid-phase thrombin, present in significant excess, is first neutralized before completing the inhibition of the surface-bound enzyme, thus increasing anticoagulation in the systemic circulation and promoting its hemorrhagic risk potential. Attempts to reduce these side effects have been directed at improving the selectivity for inhibiting thrombin in its bound state. An inhibitor with greater selectivity for surface-bound thrombin would be predicted to have a more potent antithrombotic action and reduced effects on systemic anticoagulation. See Buchanan et al, xe2x80x9cA Rationale for Targeting Antithrombotic Therapy at the Vessel Wall: Improved Antithrombotic Effect and Decreased Risk of Bleeding,xe2x80x9d Wien Klin Wochenschr (1999) 111: 81-89. This is supported by studies where the selectivity of hirudin for surface-bound thrombin was enhanced by its covalent conjugation to the anti-fibrin monoclonal antibody 59D8. See Bode et al, xe2x80x9cFibrin-Targeted Recombinant Hirudin Inhibits Fibrin Deposition on Experimental Clots More Efficiently than Recombinant Hirudin,xe2x80x9d Circulation (1994) 90(4):1956-1963; Bode et al, xe2x80x9cAntithrombotic Potency of Hirudin Is Increased in Nonhuman Primates by Fibrin Targeting,xe2x80x9d Circulation (1997) 95(4):800-804. These studies support the general concept that an increased selectively for agents or drugs that target thrombin bound to surfaces would afford a greater inhibition of intravascular/extracorporeal circuit thrombosis, enhance hemostasis in the surgical wound and potentially, decrease the duration of anticoagulant therapy. Although the above utility increases the selectivity of hirudin by its covalent attachment to fibrin- specific monoclonal antibodies, it is limited to thrombin bound to the clot and does not address improvements in the inhibition of thrombin bound to platelets, vessel wall or biomaterials which perpetuate the systemic thrombotic state.
Expression of GPIIb/IIIa receptors on the surface of activated platelets greatly enhances their adhesiveness, aggregation and adherence to the fibrin clot and the injured vessel wall. See, for example, Shen et al, xe2x80x9cInteraction of Thrombin-Activated Platelets with Extracellular Matrices (Fibronectin and Vitronectin): Comparison of the Activity of Arg-Gly-Asp-Containing Venom Peptides and Monoclonal Antibodies Against Glycoprotein IIb/IIIa Complex,xe2x80x9d J. Pharm. Pharmacol. (1997) 49(1):78-84. Thus thrombin-activated platelets promote thrombus growth indicating a need for improved thrombin inhibitors with antiplatelet therapies. See Eisenberg et al, xe2x80x9cPlatelet-Dependent and Procoagulant Mechanisms in Arterial Thrombosis,xe2x80x9d Int. J. Cardiol. (1999) 68(suppl.1):S3-S10. It is now well known that compounds that antagonize the function and/or induction of the platelet GPIIb/IIIa receptors are among the most potent antithrombotic drugs for the treatment of disease states involving platelet rich-thrombi. Indeed, these compounds inhibit platelet function or adhesion so effectively that hemorrhagic effects become a risk. See, for example, Sitges et al, xe2x80x9cMassive Pulmonary Hemorrhage in a Patient Treated with a Platelet Glycoprotein IIb/IIIa Inhibitor,xe2x80x9d Int. J. Cardiol. (1997) 62(3):269-271; Gammie et al, xe2x80x9cAbciximab and Excessive Bleeding in Patients Undergoing Emergency Cardiac Operations,xe2x80x9d C. M. Ann. Thorac. Surg. (1998) 65(2):465-469; Blankenship, xe2x80x9cBleeding Complications of Glycoprotein IIb-IIIa Receptor Inhibitors,xe2x80x9d Am. Heart J. (1999) 138(4 pt. 2):287-296. Moreover, depending on the clinical or experimental setting, these compounds have limited effects on thrombin generation and virtually no effect on thrombin activity. See Kleiman et al, xe2x80x9cInhibition of Platelet Aggregation with a Glycoprotein IIb-IIIa Antagonist Does Not Prevent Thrombin Generation in Patients Undergoing Thrombolysis for Acute Myocardial Infarction,xe2x80x9d J. Thromb. Thrombolysis (2000) 9(1):5-12; Dangas et al., xe2x80x9cEffects of Platelet Glycoprotein IIb/IIIa Inhibition with Abciximab on Thrombin Generation and Activity during Percutaneous Coronary Interventionsxe2x80x9d Am.Heart J. (1999) 138:45-54.
The combination of thrombin inhibition therapies with platelet GPIIb/IIIa receptor therapies has been recognized as desirable in the art. See PCT applications WO 99/38827 and WO 97/35592 which disclose the inclusion of hirudin, heparin and low molecular weight heparins with a platelet GPIIb/IIIa receptor antagonist. However, the inclusion of these thrombin inhibitors can significantly contribute to the overall hemorrhagic risk. Indeed, the additivity of inhibition by the combination of heparin with GPIIb/IIIa c7E3 Fab suggests these agents may have a greater bleeding liability than the use of either agent alone. See Pedicord et al., xe2x80x9cGlycoprotein IIb/IIIa Receptor Antagonists Inhibit the Development of Platelet procoagulant Activity,xe2x80x9d Thromb. Res. (1998) 90: 247-258. The therapeutic utility of lepirudin, or recombinant hirudin, is limited by its hemorrhagic potential and has shown limited benefit on thrombin generation and platelet aggregation with GPIIb/IIIa. See Koestenberger et al., xe2x80x9cEffects of the Glycoprotein IIb/IIa Receptor Antagonist c7E3 Fab and Anticoagulants on Platelet Aggregation and Thrombin potential Under High Coagulant Challenge In Vitro,xe2x80x9d Blood CoaguL (2000) 11: 425-432.
There are other problems associated with heparin and hirudin use, including antigenic reactions. Anti-hirudin antibodies are elicited in 74% of the recipients and is contraindicated in patients with a known hypersensitivity to this anticoagulant. Huhle et al, xe2x80x9cImmunologic Response to Recombinant hirudin in HIT Type II Patients during Long-Term Treatment,xe2x80x9d Br. J. Haematol. (1999) 106(1):195-201 (appendix U); Gollnick, xe2x80x9cAllergy to Heparin, Heparinoids, and Recombinant Hirudin: Diagnostic and Therapeutic Alternatives,xe2x80x9d Hautarzt (1999) 50(6):406-411. The lack of an antidote to hirudin and other thrombin inhibitors may necessitate transfusion as the only option to remedy adverse events.
Heparin-induced thrombocytopenia type II (HIT) is a consequence of heparin exposure, especially in situations such as CPB where high doses of heparin are required to manage high levels of thrombin that are continually produced during and after clinical procedures are performed. See Brister et al, xe2x80x9cThrombin Generation during Cardiac Surgery: Is Heparin the Ideal Anticoagulant?,xe2x80x9d Thromb. Haemostas. (1993) 70(2):259-262; Bauer et al, xe2x80x9cPrevalence of Heparin-Associated Antibodies without Thrombosis in Patients Undergoing Cardiopulmonary Bypass Surgery,xe2x80x9d Circulation (1997) 95:1242-1246; Pouplard et al, xe2x80x9cAntibodies to Platelet Factor 4-Heparin After Cardiopulmonary Bypass in Patients Anticoagulated with Unfractionated Heparin or a Low Molecular Weight Heparin: Clinical Implications for Heparin-Induced Thrombocytopenia,xe2x80x9d Circulation (1999) 99:2539-2536; Trossaert et al, xe2x80x9cHigh Incidence of Anti-Heparin/Platelet Factor 4 Antibodies After Cardiopulmonary Bypass surgery,xe2x80x9d Br. J. Haematol. (1998) 101(4):653-655. The bleeding problems associated with heparin use during CPB requires its neutralization by protamine salts post-CPB; this enhances the activation of inflammatory mediators, such as complement and proinflammatory cytokines which complicate outcome. See Morel et al, xe2x80x9cC5a and Thromboxane Generation Associated with Vaso-and Broncho-Constriction During Protamine Reversal of Heparin,xe2x80x9d Anesthesiology (1987) 66(5):597-604; Fehr et al, xe2x80x9cIn Vivo Complement Activation by Polyanion-Polycation Complexes: Evidence that C5a Is Generated Intravascularly During Heparin-Protamine Interaction,xe2x80x9d Clin. Immunol. Immunopathol. (1983) 29(1):7-14. Thrombin rebound (i.e., the inability of the HAT complex to neutralize trace levels of thrombin deposited on surfaces, such as the CPB circuit, surgical wound, etc.) can also occur, predisposing the patient to increased risk of thrombosis.
Accordingly, it would be desirable to be able to provide the combination of an improved thrombin inhibition therapy that provides sustained inhibition of catalytic thrombins bound to surfaces with a platelet GPIIb/IIIa receptor therapy that minimizes or prevents undesired hemorrhagic side effects, as well as potential antigenic reactions.
The present invention relates to pharmaceutical combinations that can inhibit thrombin generation and platelet aggregation with minimized or reduced hemorrhagic properties and high selectivity for surface-bound thrombin inhibition. These combinations comprise:
(a) a heparin cofactor II agonist; and
(b) a platelet glycoprotein (GP)IIb/IIIa receptor antagonist;
(c) the amount of a heparin cofactor II agonist and the amount of the platelet GPIIb/IIIa receptor antagonist combined being therapeutically effective to inhibit thrombin generation and platelet aggregation.
The present invention further relates to methods for inhibiting platelet aggregation and thrombin generation, which comprises the step of: administering (as a combined dose or as separate related doses) to a mammal in need thereof (e.g., to prevent and/or treat a variety of thrombo-embolic disorders) a combined therapeutically effective amount of a heparin cofactor II agonist and a platelet (GP) IIb/IIIa receptor antagonist.
It has been found that the administration of a combined therapeutically effective amount of a heparin cofactor II agonist or activating substance with a platelet GPIIb/IIIa receptor antagonist can provide a superior therapeutic effect in inhibiting platelet aggregation and thrombin generation (especially thrombin generation due to surface bound thrombin) than either component alone, or prior combinations of a platelet GPIIb/IIIa receptor antagonist with either unfractionated heparin, low molecular weight heparins or hirudin. Indeed, it has been found that subtherapeutic amounts of a heparin cofactor II agonist activating substance can be combined with subtherapeutic amounts of a platelet (GP) IIb/IIIa receptor antagonist to provide a therapeutically effective benefits in inhibiting platelet aggregation and thrombin generation. These surprising therapeutic benefits can be achieved while at the same time minimizing or reducing the risk of hemorrhagic side effects (e.g., prolonged bleeding), and without causing undesired antigenic responses.